The invention relates generally to methods for detecting the risk of bone fracture and, more specifically, to a method for determining the existence of low bone quality using Raman spectroscopy.
Osteoporosis is a condition that features loss of the normal density of bone and leads to abnormally porous bone. Osteoporosis affects over 44 million Americans and over one-half of these are women. The exact cause of osteoporosis is unknown. Since bones are composed of a mixture of an organic phase, primarily collagen, and an inorganic phase, primarily calcium phosphate, a prevailing thought has been that calcium deficiency leads to osteoporosis. The propensity of osteoporosis in women supports this conclusion since they often lose calcium from lactation and throughout menopause. Osteoporosis is however a poor predictor of bone fracture the clinical endpoint. The need for Bone Quality based assessments that may have improved prediction capability is well known. The current diagnostic tool, the bone mineral density scan, measures the absorption of X-rays by the calcium content in bone tissue. Accordingly, the diagnostic value of the bone density X-ray scan (DEXA scan) relies on the validity of the calcium deficiency theory of osteoporosis. Unfortunately, patients who suffer osteoporotic fractures often have normal bone density, because it is the organic phase, as well as the mineral phase, that is affected.
Bone densitometry is an involved and expensive method to measure bone density. It also has a large amount of uncertainty. The Mayo Clinic offers the following views on bone densitometry: “Bone density testing is a valuable tool in the diagnosis of osteoporosis and is a fairly accurate predictor of fracture risk. Significant differences in the various testing methods do exist, however. Central devices are more accurate, but cost significantly more than peripheral devices. The U.S. government has ordered Medicare to pay for bone density testing in the following instances: (1) If you're postmenopausal and at risk of osteoporosis; (2) if you have primary hyperparathyroidism; (3) if you have certain spinal abnormalities that might indicate a fracture; (4) if you're on long-term corticosteroid therapy, such as prednisone; or (5) if your doctor wants to assess your response to osteoporosis medications,” www.mayoclinic.com/invoke.cfm?id=WO00024
Much of the “fairly accurate predictor” language may stem from the belief that osteoporosis is a disease of the calcium content of bones. It is now, however, fairly well accepted that bone fracture risk can derive from protein problems. Bone matrix is composed of collagenous and non-collagenous bone proteins. Collagen synthesis, secretion and deposition are coordinated with that of the other matrix proteins. Cystine residues with disulphide bonding are a feature of all non-collagenous bone proteins. It is thought that the placement of cystine residues within structural proteins and consequent disulphide bonding might result in structures with varied activities. Although the mechanism of bone formation is not yet defined, the need for cystine and for some, sulfation of the bone proteins is common to all proposed mechanisms. Moreover, there is an in vivo exchange between inorganic and organic sulfate chiefly due to the synthesis and breakdown of sulfated glycosaminoglycans, which form the ground substance of bone matrix. Bone fragility may result from a disturbance, to variable degree, of these collagenous and non-collagenous bone proteins.
A publication by Akhtar and Edwards (Akhtar, W. and Edwards, H. G. M. Fourier-transform Raman spectroscopy of mammalian and avian keratotic biopolymers, Spectrochimica Acta Part A, 53, (1997) p. 81-89) outlines how mammal and avian human nails and animal beaks, quills, and claws vary in the width of the S—S band of cystine. They attribute these differences to the conformations of the S—S bond. For example, 510 cm−1 corresponds to gauche-gauche-gauche, 525 cm−1 corresponds to gauche-gauche-trans, and 540 cm−1 corresponds to trans-gauche-trans. While each of these bands can be assigned in most circumstances they are unresolved peaks or represent an unsymmetric single feature. Later Edwards used these observation to determine the health of mummified humans and birds (Edwards, A. G. M, Gniadecka, M, Ptersen. S., Hansen, J., Nielson, O., Christensen, D., Wulf, H. NIR-FT Raman spectroscopy as a diagnostic probe for mummified skin and nails, Vibrational Spectroscopy, 28, 2002, p. 3-15).
An even early publication Schrader, et al., discusses the general value of near-infrared Fourier transform Raman spectroscopy in attempting to diagnose medical conditions. This publication showed many different applications of Raman spectroscopy for medical diagnoses, but relevant to this application, they showed the spectrum of a fingernail and discussed the large S—S stretch from cystine and discussed the spectrum could indicated metabolic disorders.
Recently a group at the University of Limerick found a strong correlation between the Raman spectra of fingernail protein and osteoporosis. Moran, P., et al. Preliminary Work on the Development of a Novel Detection Method for Osteoporosis (Submitted); PCT Application No. WO 2005122893. The specific observation by the Limerick group was that the disulfide Raman feature from the amino acid cystine appears to broaden in patients with osteoporosis and increased bone fracture risk. This indicates that calcium may not be the direct culprit in osteoporosis and bone fractures and it may be a disease related to the protein component of bone material.
The problem with Limerick's approach is that it is very difficult to measure the width (more specifically the “full width at half maximum) of a peak. It is also unclear exactly what the origin of the bandwidth change is. It is possible that it is due to different conformations of the cystine in the collagen material. This will lead to more than one peak in the Raman spectrum and may manifest itself through a peak broadening.
The intensity of a Raman peak is directly related to the amount of material. If the amount of cystine is constant in healthy and osteoporotic collagen then the increased spectral width of the band should lead to a decrease in the intensity of the peal. This can be difficult to measure since natural materials such as fingernails have significant fluorescent backgrounds.